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Transglucosidase and its preparation and immobilization method

A technology for transglucosidase and BLB-28, which is applied in the field of enzyme engineering and fermentation engineering, can solve the problems of high cost of enzyme preparation, failure to realize industrialized production, and difficulty in realizing industrialized production, and achieve high enzyme stability and benefit Effect of purification and improvement of utilization rate

Active Publication Date: 2011-12-28
BAOLINGBAO BIOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] However, transglucosidase, as the most critical enzyme preparation in the production of isomaltooligosaccharides, has not been industrialized in China.
At present, the enzyme preparation is completely dependent on imports, which not only makes the key technology in the production of isomaltooligosaccharides controlled by foreign related companies, but also makes the cost of enzyme preparations high in the production of isomaltooligosaccharides
[0007] Now, in China, transglucosidase has attracted the attention of many scientific and technological workers, but on the one hand, the expression level of its transglucosidase in the wild strains screened is very low, and it is difficult to realize industrial production; on the other hand, the use of genetic engineering technology The metabolic production of Escherichia coli is generally intracellular enzymes, the extraction process is complicated, the recovery rate of enzyme activity is low, and industrial production is not used.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0060] Prepare maltose solution with pH 5.4 and 30% sugar concentration; take imported transglucosidase solution (enzyme activity is about 57-58U / ml), add 0.85L / ton of dry matter into the maltose solution, and place at 58-60°C In a water bath, keep warm for 30 hours; take a sample, put it in a boiling water bath for 15-20 minutes, inactivate the enzyme, and use liquid chromatography detection technology to analyze the product isomaltooligosaccharide components.

Embodiment 2

[0062] The aspergillus niger strain ( Aspergillus niger ) BLB-28, under the conditions of pH 6.0 and 32°C, cultured for 20 hours, activated the primary seed culture, and expanded the secondary seed cultivation; the seeds after the expanded cultivation were inoculated with an inoculation volume of 10% (V / V), and inoculated In the fermentation medium, ferment under the conditions of pH 5.5-6.0, 30-34°C, and the fermentation time is 110h. Ultrafiltration to remove fermentation bacteria and other macromolecular substances; use nanofiltration membranes with a molecular weight of 200-2000Da, concentrate by ultrafiltration at 2.0-3.0MPa, 10°C, and remove inorganic salts and small molecular substances at the same time to obtain transglucoside Enzyme solution, the enzyme activity is 60-65U.

[0063] Prepare a maltose solution with a pH of 5.4 and a sugar concentration of 30%; take the concentrated transglucosidase solution, add 0.72L / ton of dry matter to the maltose solution, place i...

Embodiment 3

[0065] The aspergillus niger strain ( Aspergillus niger ) BLB-28, under the conditions of pH 5.6 and 31°C, cultivated for 20 hours, carried out the primary seed cultivation activation, and the secondary seed expansion cultivation; the seeds after the expansion cultivation were inoculated with an inoculum volume of 8% (V / V), and inoculated In the fermentation medium, ferment under the conditions of pH 5.5-6.0, 30-35°C, and the fermentation time is 100h; Ultrafiltration to remove fermentation bacteria and other macromolecular substances; use nanofiltration membranes with a molecular weight of 200-2000Da, concentrate at 2.0-3.0MPa, 20°C, and remove inorganic salts and small molecular substances at the same time to obtain transglucoside Enzyme solution, and concentrate the enzyme solution 10 times.

[0066] Resin adsorption-sodium alginate embedding-glutaraldehyde cross-linking method immobilization. At room temperature, add 1.2kg D301(R) type resin to 1L transglucosidase soluti...

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Abstract

The invention relates to transglucosidase, its preparation method and a immobilization method, which belong to the fields of enzyme engineering and fermentation engineering. More specifically, the invention provides the following processes: 1) Aspergillus niger BLB-16 is screened from soil as an original strain, processes of mutation treatment, optimization and screening are performed to obtain an optimized bacterial strain (BLB-28) for fermenting; 2) a fermentation broth is carried out a heat sterilization to obtain transglucosidase liquid; 3) transglucosidase liquid is performed a nanofiltration to concentrate; 4) processes of resin adsorption, sodium alginate entrapment, immobilization by a glutaraldehyde cross-linking method are carried out for preparing immobilized enzymes. The prepared transglucosidase is suitable for an application in the industrial fields such as foodstuff, medicine, feed and the like, and used for producing isomaltose hypgather.

Description

technical field [0001] The invention belongs to the fields of enzyme engineering and fermentation engineering, and relates to a transglucosidase and a preparation and immobilization method thereof. Specifically, the Aspergillus niger strain screened from soil ( Aspergillus niger ) BLB-16 is the starting strain, the optimized strain BLB-28 is obtained through mutagenesis and screening, and the transglucosidase solution is prepared through fermentation, sterilization and concentration, and the immobilization technology of the transglucosidase is also involved. Background technique [0002] Transglucosidase (Transglucosidase), also known as glucosidase, α-glucosidase, α-transglucosidase, α-D-glucosidase, etc., according to "Enzyme Nomenclature and International Classification Number", Belongs to transferases, specifically EC 2.4.1.24, CAS 9030-12-0. [0003] According to the classification standard of "Hygienic Standards for the Use of Food Additives" (GB2760), transglucosida...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14C12N9/10C12N11/10C12N11/04C12P19/18C12R1/685
Inventor 刘宗利王乃强袁卫涛王彩梅李方华
Owner BAOLINGBAO BIOLOGY
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